NAHEMS National Animal Health Emergency Management System

Transcription

1 NAHEMS GUIDELINES: VACCINATION FOR CONTAGIOUS DISEASES APPENDIX B: VACCINATION FOR CLASSICAL SWINE FEVER FAD PReP Foreign Animal Disease Preparedness & Response Plan NAHEMS National Animal Health Emergency Management System United States Department of Agriculture Animal and Plant Health Inspection Service Veterinary Services AUGUST 2017

2 The Foreign Animal Disease Preparedness and Response Plan (FAD PReP)/National Animal Health Emergency Management System (NAHEMS) Guidelines provide a framework for use in dealing with an animal health emergency in the United States. This FAD PReP/NAHEMS Guidelines was produced by the Center for Food Security and Public Health, Iowa State University of Science and Technology (ISU), College of Veterinary Medicine, in collaboration with the U.S. Department of Agriculture (USDA) Animal and Plant Health Inspection Service through a cooperative agreement. This Guidelines document has undergone review by USDA Legislative and Public Affairs. This FAD PReP/NAHEMS Guidelines reflects updates to the 2012 version, completed in August Please send questions or comments to: Center for Food Security and Public Health National Preparedness and Incident Coordination 2160 Veterinary Medicine Animal and Plant Health Inspection Service Iowa State University of Science and Technology U.S. Department of Agriculture Ames, IA River Road, Unit 41 Phone: /Fax: Riverdale, Maryland cfsph@iastate.edu, FAD.PReP.Comments@aphis.usda.gov Subject line: FAD PReP/NAHEMS Guidelines While best efforts have been used in developing and preparing the FAD PReP/NAHEMS Guidelines, the U.S. Government, USDA, ISU and other parties, such as employees and contractors contributing to this document, neither warrant nor assume any legal liability or responsibility for the accuracy, completeness, or usefulness of any information or procedure disclosed. The primary purpose of these FAD PReP/NAHEMS Guidelines is to provide guidance to those government officials responding to a foreign animal disease outbreak. It is only posted for public access as a reference. The FAD PReP/NAHEMS Guidelines may refer to links to various other Federal and State agencies and private organizations. These links are maintained solely for the user's information and convenience. If you link to such site, please be aware that you are then subject to the policies of that site. In addition, please note that USDA does not control and cannot guarantee the relevance, timeliness, or accuracy of these outside materials. Further, the inclusion of links or pointers to particular items in hypertext is not intended to reflect their importance, nor is it intended to constitute approval or endorsement of any views expressed, or products or services offered, on these outside web sites, or the organizations sponsoring the web sites. Trade names are used solely for the purpose of providing specific information. Mention of a trade name does not constitute a guarantee or warranty of the product by USDA or an endorsement over other products not mentioned. USDA prohibits discrimination in all its programs and activities on the basis of race, color, national origin, sex, religion, age, disability, political beliefs, sexual orientation, or marital or family status. (Not all prohibited bases apply to all programs.) Persons with disabilities who require alternative means for communication of program information (Braille, large print, audiotape, etc.) should contact USDA s TARGET Center at (202) (voice and telecommunications device for the deaf [TDD]). To file a complaint of discrimination, write USDA, Director, Office of Civil Rights, Room 326-W, Whitten Building, 1400 Independence Avenue SW, Washington, DC or call (202) (voice and TDD). USDA is an equal opportunity provider and employer. ISU does not discriminate on the basis of race, color, age, ethnicity, religion, national origin, pregnancy, sexual orientation, gender identity, genetic information, sex, marital status, disability, or status as a U.S. veteran. Inquiries regarding non-discrimination policies may be directed to Margo Foreman, Director of Equal Opportunity, 3350 Beardshear Hall, Ames, Iowa 50011, Tel , eooffice@iastate.edu. i

3 THE IMPERATIVE FOR FOREIGN ANIMAL DISEASE PREPAREDNESS AND RESPONSE Why Foreign Animal Diseases Matter Preparing for and responding to foreign animal diseases (FADs) such as highly pathogenic avian influenza (HPAI) and foot-and-mouth disease (FMD) are critical actions to safeguard the nation s animal health, food system, public health, environment, and economy. FAD PReP, or the Foreign Animal Disease Preparedness and Response Plan, prepares for such events and provides guidance for activities during a response. Since 2014, three HPAI outbreaks in the United States have cost over $880 million, just for indemnity payments and response activities on premises. Studies have estimated a likely national welfare loss between $ billion 1 for an FMD outbreak in California, depending on delay in diagnosing the disease. 2 The economic impact of an FAD outbreak results from lost international trade and disrupted interstate trade, as well as from costs directly associated with the eradication effort, such as depopulation, indemnity, disposal, and virus elimination. In addition, there are direct and indirect costs related to foregone production, unemployment, and losses in related businesses. The social and psychological impact on owners and growers can be significant. Diseases with zoonotic potential, such as HPAI and Nipah/Hendra, may also pose a threat to public health. Challenges of Responding to an FAD Event Responding to an FAD event large or small is complex and difficult, challenging all stakeholders involved. Response activities require significant prior preparation. There are imminent and problematic disruptions to interstate commerce and international trade. A response effort must have the capability to be rapidly scaled up or down according to the needs of the specific incident. This involves many personnel, resources, and possibly veterinary countermeasures. Not all emergency responders have specific food and agriculture skills required in areas such as biosecurity, quarantine and movement control, epidemiological investigation, diagnostic testing, depopulation, disposal, and possibly emergency vaccination. Establishing widely communicated and understood response goals and guidelines, as accomplished by the FAD PReP materials, helps to broaden awareness of common objectives as well as potential problems. 1 Carpenter TE, O Brien JM, Hagerman AD, & McCarl BA Epidemic and economic impacts of delayed detection of foot-andmouth disease: a case study of a simulated outbreak in California. J Vet Diagn Invest. 23: Estimates based on models may vary: Ekboir (1999) estimated a loss of between $8.5 and $13.5 billion for an FMD outbreak in California. Ekboir JM Potential Impact of Foot-and-Mouth Disease in California: the Role and Contribution of Animal Health Surveillance and Monitoring Services. Agricultural Issues Center. University of California, Davis. ii

4 Lessons Learned from Past FAD Outbreaks The foundation of FAD PReP is the lessons learned from past FAD incidents. FAD PReP is based on the following: Achieving rapid FAD detection and tracing. Providing processes for emergency planning that respect local knowledge. Integrating State-Federal-Tribal-industry planning processes. Ensuring that there are clearly defined, obtainable, and unified goals for response. Having a unified Incident Command that can act with speed and certainty. Employing science- and risk-based management approaches to an FAD response. Ensuring that all guidelines, strategies, and procedures are communicated effectively to responders and stakeholders. Identifying trained personnel and resources that are required for an effective incident response. Trying to resolve competing interests prior to an outbreak and addressing them quickly during an outbreak. FAD PReP Mission and Goals The mission of FAD PReP is to raise awareness, expectations, and develop capabilities surrounding FAD preparedness and response. The goal of FAD PReP is to integrate, synchronize, and deconflict preparedness and response capabilities as much as possible before an outbreak by providing goals, guidelines, strategies, and procedures that are clear, comprehensive, easily readable, easily updated, and that comply with the National Incident Management System. In the event of an FAD outbreak, the three key response goals are to: (1) detect, control, and contain the FAD in animals as quickly as possible; (2) eradicate the FAD using strategies that seek to stabilize animal agriculture, the food supply, the economy, and to protect public health and the environment; and (3) provide science- and risk-based approaches and systems to facilitate continuity of business for noninfected animals and non-contaminated animal products. Achieving these three goals will allow individual livestock facilities, States, Tribes, regions, and industries to resume normal production as quickly as possible. They will also allow the United States to regain FAD-free status without the response effort causing more disruption and damage than the disease outbreak itself. FAD PReP Documents and Materials FAD PReP is not just one, standalone FAD plan. Instead, it is a comprehensive U.S. preparedness and response strategy for FAD threats, both zoonotic and non-zoonotic. The following section provides examples of the different types of FAD PReP documents available. Strategic Plans Concept of Operations APHIS Foreign Animal Disease Framework: Roles and Coordination (FAD PReP Manual 1-0): This document provides an overall concept of operations for FAD preparedness and response for APHIS, explaining the framework of existing approaches, systems, and relationships. APHIS Foreign Animal Disease Framework: Response Strategies (FAD PReP Manual 2-0): This document provides significant detail on response strategies that will be conducted in an FAD outbreak. APHIS Foreign Animal Disease Framework: Information Management & Reporting (FAD PReP Manual 3-0): This document explains how information is managed and reported in FAD incidents. iii

5 FAD Investigation Manual (FAD PReP Manual 4-0): This field-ready manual provides detailed information on completing an FAD investigation from start to finish. A Partial List of FAD Stakeholders (FAD PReP Manual 5-0): This guide identifies key stakeholders with whom the National Preparedness and Incident Coordination (NPIC) Center collaborates. NAHEMS Guidelines These documents describe many of the critical preparedness and response activities, and can be considered as a competent veterinary authority for responders, planners, and policy-makers. Industry Manuals These manuals describe the complexity of industry to emergency planners and responders and provide industry a window into emergency response. Disease Response Plans Response plans are intended to provide disease-specific information about response strategies. They offer guidance to all stakeholders on capabilities and critical activities that would be required to respond to an FAD outbreak. Standard Operating Procedures (SOPs) for Critical Activities For planners and responders, these SOPs provide details for conducting critical activities such as disposal, depopulation, cleaning and disinfection, and biosecurity that are essential to effective preparedness and response to an FAD outbreak. These SOPs provide operational details that are not discussed in depth in strategy documents or disease-specific response plans. Continuity of Business Plans (commodity-specific plans developed by public-private-academic partnerships) Known as the Secure Food Supply Plans, these materials use science- and risk-based information to facilitate market continuity for specific products in an outbreak. APHIS Emergency Management APHIS Directives and Veterinary Services (VS) Memorandums provide important emergency management policy. These documents provide guidance on topics ranging from emergency mobilization, to FAD investigations, to protecting personnel from HPAI. Most of these documents are available publicly, at iv

6 PREFACE The Foreign Animal Disease Preparedness and Response Plan (FAD PReP)/National Animal Health Emergency Management System (NAHEMS) Guidelines provide the foundation for a coordinated national, regional, state and local response in an emergency. As such, they are meant to complement non- Federal preparedness activities. These guidelines may be integrated into the preparedness plans of other Federal agencies, State and local agencies, Tribal Nations, and additional groups involved in animal health emergency management activities. This Appendix B: Vaccination for Classical Swine Fever is a supplement to FAD PReP/NAHEMS Guidelines: Vaccination for Contagious Diseases, and covers the disease-specific strategies and general considerations of vaccination. Both documents are components of APHIS FAD PReP/NAHEMS Guideline Series, and are designed for use by APHIS Veterinary Services (VS), and other official response personnel in the event of an animal health emergency, such as the natural occurrence or intentional introduction of a highly contagious foreign animal disease in the United States. Appendix B: Vaccination for Classical Swine Fever, together with the Vaccination for Contagious Diseases Guidelines, provide guidance for USDA employees on principles of vaccination for classical swine fever for animal health emergency deployments. This Appendix B: Vaccination for Classical Swine Fever provides information for incident management personnel and other responders associated with vaccination activities. The general principles discussed in this document are intended to serve as a basis for understanding and making sound decisions regarding vaccination in a classical swine fever emergency. As always, it is important to evaluate each situation and adjust procedures to the risks present in the situation. The FAD PReP/NAHEMS Guidelines are designed for use as a preparedness resource rather than as a comprehensive response document. v

7 APHIS DOCUMENTS Key APHIS documents complement this Appendix B: Vaccination for Classical Swine Fever, Strategies and Considerations and provide further details when necessary. This document references the following APHIS documents: APHIS Foreign Animal Disease Framework documents o Roles and Coordination (FAD PReP Manual 1-0) o Response Strategies (FAD PReP Manual 2-0) Classical Swine Fever Response Plan (The Red Book) These documents are available on the FAD PReP collaboration website at: For the full listing of all references, including other APHIS documents, see section 19. References. vi

8 Summaries of each section can be accessed from the table of contents and are followed by more detailed descriptions of the material. 1. Purpose Background Overview of CSF... 2 Summary Serotypes and Strains Species Affected Pathogenesis Persistent Infection Clinical Signs Acute Infection Subacute Infection Chronic Infection Persistent Infection Transmission Incubation Transmission Routes Survival Vaccination and Virus Transmission Detection of Infected Animals... 6 Summary Detecting Infected Animals by Identifying Virus, Nucleic Acids, or Antigen Identification of the Virus Identification of Viral Antigen Detecting Infected Animals by Serological Assays Virus Neutralization Tests (VNTs) Antibody Detection ELISAs Serological Assays in Development The Use of Diagnostic Tests in Outbreaks Outbreak Testing: Real World Examples CSF Vaccines...11 Summary Types of CSF Vaccines Live Attenuated Virus (LAV) Vaccines E2 Marker Vaccines Pestivirus Chimeric Vaccines Additional Subunit Vaccines/Immunogenic Peptides DNA Vaccines Viral Vector Vaccines Trans-Complemented Deletion Mutants (Replicons) Additional Approaches Production of CSF Vaccines LAV Vaccines vii

9 5.2.2 Vaccines Produced Through Biotechnology Vaccine Banks National Veterinary Stockpile International Examples CSF Vaccines from Commercial Manufacturers Vaccine Licensing Vaccine Matching, Efficacy, and Safety...16 Summary Vaccine Matching Vaccine Efficacy and Effectiveness Vaccine Safety Effects of Vaccination on Virus Transmission Examples of R Values for CSFV Vaccines Onset of Protective immunity Duration of Immunity Maternal Antibodies Vaccine Withdrawal Times in Meat Strategies for Vaccine Use...19 Summary CSF Vaccination Strategies in the U.S CSF Vaccination Strategies in the EU Vaccination Terminology and CSF Applications Prophylactic Vaccination Emergency Vaccination Protective Emergency Vaccination Suppressive (or Damping Down ) Emergency Vaccination Targeted Vaccination Ring Vaccination Barrier Vaccination Blanket Vaccination Establishing a Vaccination Zone Advantages and Disadvantages of CSF Vaccination Field Experiences with CSF Vaccination...22 Summary Brazil Bulgaria Germany Great Britain Israel Mexico Netherlands Republic of Korea Romania United States Modeling Studies and Vaccination Movement Restrictions and Vaccination Permanent Identification of Vaccinated Animals Logistic and Economic Considerations...28 viii

10 Summary Technical Feasibility of Vaccination Epidemiological Considerations Weather Distance Swine Density Feral Swine Infection with Other Pathogens Costs Associated with Vaccination Vaccination and Market Effects Effect of Vaccination on OIE Status Vaccination of Special Populations Public Acceptability of Vaccination as a Component of CSF Eradication...31 Summary Classical Swine Fever Disease as a Zoonosis The Use of Meat from Vaccinated and/or Potentially Infected Animals Procedures to Inactivate CSFV in Animal Products Procedures for Marketing Animal Products After Emergency Vaccination Public Acceptability of Other CSF Control Strategies References Acknowledgements...44 Glossary...46 Acronyms...50 ix

11 1. PURPOSE This Appendix is intended to provide relevant information for Federal and State officials, and other interested parties, who will participate in decision-making related to vaccine use in an outbreak of classical swine fever (CSF) in the United States (U.S.). The following topics are presented and discussed: Important characteristics of CSF; Characteristics of vaccines; Strategies for vaccine use; and Various factors that must be considered when designing an effective vaccination program. 2. BACKGROUND CSF (also known as hog cholera) is a highly contagious viral disease of swine. The U.S. eradicated its last case of CSF in August However, a possible disease re-introduction continually threatens the U.S. swine herd. CSF virus (CSFV) could enter the U.S. through multiple routes. The intentional release of CSFV into the U.S. swine herd is a real concern. Unintentional introduction is also possible. Employees and owners of hog production systems, who travel all over the world, as well as visitors who arrive from countries with endemic CSF, could unintentionally expose pigs to the virus. The clinical signs associated with CSF resemble many endemic diseases, which may delay diagnosis and make control even more difficult. Industry estimates place over one million swine in trucks on the road every day. 2 During transport, any pig exposed to CSFV would have the potential to spread the disease to another location before it is diagnosed. Artificial insemination (AI) is a technology which has greatly benefitted the U.S. swine industry; however, if a boar stud becomes infected with CSFV, infected semen could be distributed throughout the country unknowingly. 3 Feral swine are now found throughout most of the southern U.S. and their range is ever expanding. 4 CSF would be even more difficult to eradicate from the U.S. if it was also found in feral swine. An appropriate and usable CSF response plan must be in place before a diagnosis is confirmed to enable an effective swine industry response. CSF is endemic in many parts of the world. CSF is found in some areas of Asia, Africa, South and Central America, and the Caribbean islands. 5 Nearby threats specifically include Haiti, the Dominican Republic, and Cuba where CSF has been found as recently as late 2016 and Mexico, where the last case was reported in mid CSF has been eradicated from the U.S., Canada, New Zealand, Australia, and from domestic swine operations in most of western and central Europe. Outbreaks in CSF-free countries have resulted in CSF infection on multiple farms and significant economic losses for swine industries in those countries. In 1994, Germany reported 117 farms infected, and Belgium reported 48 CSFV-positive farms. 7 Modeling of an outbreak that occurred from , involving 429 farms in the Netherlands, showed an estimated $423 million in losses for swine farmers and $596 million in losses for related industries. 7 Terpstra et al. 8 estimated greater losses in the Netherlands for the same outbreak totaling over $2 billion (direct losses only). Paarlberg et al. 9 calculated potential economic losses caused by a hypothetical CSF outbreak in the U.S. Eleven million hogs were FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 1

12 destroyed in this scenario, resulting in estimated losses ranging from $2.6 billion to $4.1 billion when considering the value of destroyed animals, the effect on breeding herd numbers, product demand, and effect on exports. Controlling CSF in areas that are pig-dense has proven to be very challenging. Although the measures of stop animal movement, isolation, and stamping-out helped to control the outbreak in the Netherlands, it was at a great economic loss. 10 Measures to control CSF outbreaks in the Netherlands, England, and Belgium did not include CSF vaccination. For countries of the European Union (EU), utilizing CSF vaccination is prohibited unless the affected country requests and is granted permission to carry out emergency vaccination in addition to control measures already underway, according to Article 19 of EU Directive 2001/89/EC. 11 For example, beginning in 2006, Romania determined that in order to eradicate CSF, vaccination would be required. Contingency plans, including the use of CSF vaccine, were submitted by Romania to the Commission on November 9, 2006 for the control of CSF and approved under Directive 2007/19/EC. 12 Stop movement orders could prevent or slow CSF spread during an outbreak, but may compromise animal welfare. With the management practices of the U.S. swine industry, many animals remain on a site until a specified weight or age. For example, pigs may be placed in a nursery from weaning, at about 3 weeks of age, until they reach about 50 pounds body weight. At that time, they are moved into a finishing building. If a stop movement were in place, animals in a nursery would continue to grow and become overcrowded. In addition, young animals that need to be weaned could not be transported to a new site or building if it has not been emptied. According to Pluimers et al., 10 during the CSF outbreak in the Netherlands during , the welfare of the pigs during a stop movement was a concern. As animals became overcrowded and pigs began to suffer health problems, authorities implemented a buyout plan and carcasses were destroyed. In 2004, participants in the World Organization for Animal Health (OIE) International Conference on the Control of Infectious Animal Diseases by Vaccination concluded that mass slaughter is no longer acceptable as the main technique for disease control and eradication due to ethical, ecological, and economic concerns. 13 They recommended that methods for disease prevention, control, and eradication be reviewed, and advised an increased emphasis on vaccination. When initial control measures such as stamping-out, quarantine, and stop movement do not contain a CSF outbreak, the use of vaccine must be considered. According to DeHaven, 14 The decision to use, or not to use, a vaccine in the face of a foreign animal disease outbreak can be complex and have far-reaching socio-economic consequences. Incorrect decisions or delays occurring during the actual outbreak can be costly. Factors that can influence the decision to vaccinate include the number of herds affected, how quickly the disease is spreading, personnel available to assist in the response effort, and the number of feral swine in the area. 3. OVERVIEW OF CSF Summary CSFV is a member of the genus Pestivirus and family Flaviviridae. The small, enveloped, single stranded RNA virus is closely related to ruminant pestiviruses that cause bovine viral diarrhea (BVD) and border disease of sheep. In recent years, atypical pestiviruses have been identified in several species including pigs. Only one CSF serotype exists; however, viral strains can be divided into three genotypes with three to four sub-genotypes that show a distinct geographical pattern. All species of domestic pigs (Sus scrofa domesticus), feral pigs, and wild pigs including European wild boar (Sus scrofa scrofa) and collared peccaries are thought to be susceptible to CSFV infection. FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 2

13 Humans and other livestock species do not appear to be affected by CSF. Virus shedding can begin before the onset of clinical signs, and occurs throughout the course of acute or subclinical disease. Chronically or persistently infected pigs can shed virus continuously or intermittently for months. Sows can be infected with CSFV at any stage of gestation, and the virus can then cross the placenta and infect the fetuses. The outcome of prenatal infection depends on strain virulence and the time of gestation at which infection occurs. Sows infected with CSFV during gestation may deliver stillborn, aborted, or mummified pigs. Pigs born alive may be persistently infected. Persistently infected pigs may appear asymptomatic at birth; however, congenital tremors may develop and stunting may become apparent over time. Persistently infected piglets may survive to 6 months, rarely up to a year, while shedding the virus and acting as a source of infection for other pigs. Cerebellar hypoplasia is evident more frequently in pigs born to sows infected with CSFV prior to 43 days of gestation. Clinical signs vary depending on the stage of infection, type of disease (acute, subacute, chronic or persistent/late onset), and virulence of the strain. Clinical signs associated with acute infection with a highly virulent strain include a high fever, huddling, weakness, drowsiness, anorexia, conjunctivitis, and constipation followed by diarrhea. About 2 4 weeks after infection, purple discoloration of the skin on the abdomen, inner thighs, or ears may be visible, or hemorrhages may be evident. Vomiting bile may occur, or respiratory signs may develop. Some pigs show neurologic signs such as incoordination or unsteadiness, which may progress to posterior paresis or convulsions in the terminal stages. Clinical signs are generally less severe with the subacute form due to infection with lower virulence strains. Pigs that are chronically infected show mostly nonspecific clinical signs; weight loss can occur over time. Transmission between pigs occurs mainly by the oral or oronasal routes via direct or indirect contact. Virus can be shed in saliva, lacrimal secretions, blood, urine, feces, and semen. Transmission may occur through ingestion of uncooked garbage containing infected pork products. Genital transmission and transmission via AI also occurs. CSFV can be transmitted on fomites and by mechanical vectors. Airborne transmission seems to be possible over short distances; however, the maximum distance the virus can travel is unclear. The incubation period for acute disease can range from 2 to 14 days, depending on the virulence of the strain, the route of infection, and the dose. CSFV is easily transmitted due to its ability to persist in the environment and in pork products. CSFV can survive in chilled pork up to 3 months, in frozen pork and pork products up to 4 years, and in salted or smoked meat up to 180 days. 3.1 Serotypes and Strains CSFV is a member of the genus Pestivirus and family Flaviviridae. 15 The small, enveloped, single stranded RNA virus is closely related to the ruminant pestiviruses that cause BVD and border disease of sheep. 16 In recent years, atypical pestiviruses have been identified in several species including pigs Only one CSF serotype exists; however, viral strains can be divided into three genotypes with three to four sub-genotypes that show a distinct geographical pattern. 26 The viral particle is composed of four FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 3

14 structural proteins: the core protein (protein C) and envelope glycoproteins Erns, E1, and E2. 27 CSFV strains can vary considerably in virulence. 3.2 Species Affected All species of domestic pigs (Sus scrofa domesticus), feral pigs, and wild pigs including European wild boar (Sus scrofa scrofa) and collared peccaries are thought to be susceptible to CSFV infection. Humans and other livestock species do not appear to be affected by CSF. 3.3 Pathogenesis The most common form of CSFV transmission in pigs is oronasal. 28 Following intranasal inoculation with CSFV, the virus replicates primarily in the tonsils before spreading to other lymphoid organs, including regional lymph nodes, and then the peripheral blood, bone marrow, and visceral lymph nodes. 28 Spread of the virus within the animal usually occurs in 5 6 days. 32 Virus shedding can begin before the onset of clinical signs, and occurs throughout the course of acute or subclinical disease. 33 Chronically or persistently infected pigs can shed virus continuously or intermittently for months. 34, Persistent Infection Infection with CSFV during pregnancy leads to persistent infection where affected pigs do not mount an adequate immune response. 26 Persistent infection is sometimes referred to as the prenatal course or late onset CSF. Experimentally, when sows were infected with CSFV on either day 22 or 43 of gestation, pigs born showed a variety of clinical signs including tremors. 36 Of those with tremors, 83% of those pigs had cerebellar hypoplasia. Several piglets died within a few days of birth. Pigs from sows infected after day 72 of gestation did not exhibit severe tremors, although a majority of these piglets were either mummified or stillborn. 36 Tremors became less evident as pigs grew older and continued to shed virus. Van Oirschot et al. 37 produced different results after infecting 4 sows at days 40, 65, and 90 of gestation with a low virulence CSFV strain. Transplacental transmission did not occur in 2/4 sows infected at 40 days gestation and 2/4 sows infected at 90 days gestation. Two different sows infected at 40 days gestation gave birth to pigs which all tested positive for virus at birth, and 1/4 sows infected at 65 days gestation also gave birth to pigs all of which tested positive at birth. 37 Concerning persistently infected animals, it was concluded from this experiment that sows infected with a low virulence strain of CSFV, at an earlier stage of gestation, will produce a greater number of persistently infected pigs. 37 Dahle et al. 38 inoculated sows between 70 and 90 days gestation and observed persistently infected pigs born to these sows. Persistent infection has also been experimentally induced by infecting newborn piglets within 8 and 48 hours after birth. 39, Clinical Signs Pigs infected with CSFV may show a variety of clinical signs depending on the stage and type of infection (acute, subacute, chronic or persistently infected) and virulence of the strain Acute Infection Clinical signs associated with acute infection include high fever, huddling, weakness, drowsiness, anorexia, conjunctivitis, and constipation followed by diarrhea. 28,41,42 Approximately 2 4 weeks later, purple discoloration of the skin on the abdomen, inner thighs, or ears may be visible, or hemorrhages may be evident. Vomiting bile may occur, or respiratory signs may develop. Pigs may show neurologic signs such as incoordination or unsteadiness, which may progress to posterior paresis or convulsions in the terminal stages. 5,42 Secondary infections can complicate the clinical diagnosis. 26 If bloodwork is conducted, severe leukopenia is usually seen. Pigs with acute CSF often die within days postinfection Subacute Infection FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 4

15 Moderately virulent strains of CSFV can cause subacute disease in which the clinical signs are less severe. However, fever may persist for 2 3 weeks. Survival of pigs with subacute CSF varies as some survive for longer periods, while others die within a month Chronic Infection Chronic infection is linked to CSFV strains of moderate virulence. 34,43 When the immune system cannot eliminate the virus, pigs develop the chronic form of CSF. Nonspecific clinical signs (e.g., fever, depression, wasting, and diffuse dermatitis) are most commonly seen. 26 Chronically infected pigs lose weight as severe lesions develop in the ileum and rectum. 42 Chronically infected pigs can shed CSFV for 1 3 months. 44 Pigs developing the chronic form of CSF may survive 2 3 months before they die Persistent Infection Sows can be infected with CSFV at any stage of gestation, and the virus can then cross the placenta and infect the fetuses leading to persistent infection. Persistent infection is sometimes referred to as the prenatal course or late onset CSF. The outcome of prenatal infection will depend on the virulence of the strain and the time of gestation. 42 If infected in early pregnancy with a strain of moderate or low virulence, pigs may be aborted, stillborn, or mummified. Infection of the sow around days of gestation may result in persistently infected pigs, depending on the strain virulence. 42 When persistently infected pigs are born alive, some develop a congenital tremor while others are asymptomatic at birth. 37,42 Persistently infected animals may not show signs, such as stunted growth, for several months following birth. 42,45 Some pigs will survive for more than 6 months, but rarely past one year, while shedding the virus and potentially spreading the disease. 35 The differential diagnosis for CSF includes both endemic and foreign animal diseases such as African swine fever, salmonellosis, porcine dermatitis and nephropathy syndrome, erysipelas, porcine circovirusassociated disease, hemolytic disease of the newborn, porcine reproductive and respiratory syndrome, pasteurellosis, actinobacillosis, Haemophilus suis infection, thrombocytopenic purpura, anticoagulant (e.g. warfarin) poisoning, salt poisoning, pseudorabies, parvovirus infection, and eperythrozoonosis. 5, Transmission Incubation The OIE listed incubation period for CSF ranges from 2 to 14 days. 46 Pasick reports 3 to 4 days may be typical. 47 Chronic infection may not appear until up to 3 months following virus exposure. 46 Under field conditions, the disease may not be diagnosed in a herd for weeks as the clinical signs resemble domestic diseases. For example, in the Netherlands, on January 15, 1997, a practitioner observed atypical clinical signs in finishing pigs. 48 He suspected pneumonia and prescribed antibiotics. When the pigs did not respond to treatment, the practitioner suspected porcine reproductive and respiratory syndrome (PRRS) and submitted two pigs to the diagnostic laboratory on January 21. The laboratory diagnosed CSF on February 4, 1997, more than 2 weeks after the pigs began to show clinical signs Transmission Routes CSF is highly contagious. Virus can be shed in saliva, lacrimal secretions, blood, urine, feces and semen. 3,37,49-51 Transmission between pigs occurs mainly by the oral or oronasal routes via direct or indirect contact. 28,42,52,53 Some experts regard direct contact as the most important route of CSF transmission. 54 Feral swine roam in many states throughout the U.S. If CSFV were to infect feral swine, the health of the U.S. domestic swine herd would be threatened. In Germany, direct or indirect contact with CSFV-infected wild boar was found to be the cause of CSFV transmission to domestic swine. 53 CSFV can also be spread by genital transmission or AI as boar semen may contain the virus. 3 In the CSF outbreak in the Netherlands, two AI studs became infected. Because the boar studs were FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 5

16 allowed to continue shipping semen, they were suspected of potentially infecting 21 sow herds. 8 Infected carrier sows may give birth to persistently infected pigs. Pigs can acquire CSFV through ingestion of uncooked garbage containing infected pork products. 53 Pork products may be smuggled into the U.S. from countries with endemic CSF, enabling long-distance transmission of CSF. Garbage feeding has been suspected in other countries as a means of CSF introduction. For example, in Bulgaria, non-vaccinated pigs fed uncooked table scraps tested positive for CSFV in March CSFV can survive in chilled pork up to 3 months, in frozen pork and pork products up to 4 years, 59 and in salted or smoked meat up to 180 days. 5,57,59-62 According to Kleiboeker, 63 a hiker feeding part of a ham sandwich to a sow herd may have introduced CSFV into the United Kingdom in Fomites and mechanical vectors can also play a role in CSFV transmission. Dorset et al. 64 reported that stable flies and house flies can transmit CSFV from sick pigs to healthy pigs. It is still unclear if birds play a role in CSF transmission. 54 The role of cats, dogs, and rodents in CSF transmission has also been questioned. Research by Dewulf et al. 65 showed that cats, dogs, and rats do not serve as reservoirs of CSFV; however, mechanical transmission may still be possible. Dewulf et al. 66 demonstrated that airborne transmission is possible under experimental conditions, although the maximum distance the virus can spread is unclear. While aerosol transmission was documented only within a radius of 250 meters in some studies, transmission occurred up to 1 km in another. 54 Fomites such as transportation vehicles pose a threat in transmitting CSFV when not properly cleaned and disinfected. Although it has not been proven, transportation vehicles originating from Germany are thought to have introduced CSFV into the Netherlands during the CSFV outbreak. 48 The very cold weather made properly cleaning and disinfecting of the transport vehicles difficult Survival Estimates of CSFV survival in pens and on fomites under field conditions vary. Weesendorp et al. 67 found that CSFV was no longer detectable in the feces after 42 days (when pigs were infected with the moderately virulent Paderborn strain) and after 64 days (when pigs were infected with the highly virulent Brescia strain). Neither strain of the virus could be detected in urine after 18 days post infection. 67 While initial concentrations and strain of the virus in feces will affect the survival time, at 20 C (68 F), the virus was inactivated in feces within 3 (Paderborn strain) to 5 days (Brescia strain) and in hours at 30 C (86 F). 67 Information on CSFV survival in animal products can be found in section Vaccination and Virus Transmission Effective vaccination can decrease transmission between animals by 1) decreasing the susceptibility of animals to infection, and 2) reducing virus shedding, if a vaccinated animal becomes infected. 4. DETECTION OF INFECTED ANIMALS FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 6

17 Summary Diagnosis of CSF based on clinical signs alone is almost impossible. Several OIE-approved tests are available; preferred diagnostic samples include serum (for live animals) or tissues such as tonsil and ileum (for dead animals). Diagnostic tests that detect virus, viral nucleic acid, or viral antigen include virus isolation, reverse transcription polymerase chain reaction (RT-PCR), real-time (or quantitative) PCR (rrt-pcr or qrt- PCR), direct immunofluorescence (fluorescent antibody test, FAT), immunoperoxidase staining, and antigen-capture enzyme linked immunosorbent assay (ELISA). Virus isolation is the test of choice to confirm CSFV infection; however, it is slow and labor intensive. After growing virus in PK-15 cells (or another porcine line), cultures are examined by FAT after 24 to 72 hours, or in 4- to 5-day-old cultures by immunoperoxidase staining. Samples shipped for virus isolation should be refrigerated but not frozen. RT-PCR is a fast and sensitive diagnostic method that can detect CSF nucleic acids in pigs throughout the course of disease, from the preclinical stage to recovery. Blood samples from live pigs or tissues samples collected during necropsy can be tested with results in 48 hours or less. CSFV antigen can be rapidly detected using the FAT. This test uses frozen tissue sections which are stained directly with an anti-csf immunoglobulin conjugated to a fluorescence marker, or indirectly with a fluorescent conjugate, and examined by fluorescence microscopy. 45 Detecting CSF virus in the tonsil is more likely 2 to 15 days post infection when animals are showing clinical signs of CSF. Vaccine administration and infection with ruminant pestiviruses may affect FAT results in some cases. Immunoperoxidase staining is a test that uses monoclonal antibodies (MAbs) and can differentiate field strains of CSFV, vaccine strains of CSFV, and other pestiviruses including those from ruminants. Antigen-capture ELISA is a good tool for early diagnosis of CSFV at the herd level; however, sensitivity is low and the assay is not appropriate for CSFV surveillance purposes or detection of CSFV in individual animals. Serology is used for diagnosis and surveillance. Antibodies cannot be detected until at least 21 days post infection and persist for life. Serological tests include virus neutralization (neutralizing peroxidase-linked assay, fluorescent antibody virus neutralization test, comparative neutralization test) and various ELISAs. Diagnostic tests with the ability to detect infection in vaccinated animals (DIVA) have been developed, but to date, most are useful only at the herd level because of less than optimal sensitivity and specificity. Many countries, including those in the EU, accept the use of rrt-pcr for screening and confirmation of suspected cases of CSF. However, a positive result must always be confirmed by another test such as virus isolation. Diagnosis of CSF based on clinical signs alone is almost impossible. The clinical presentation of CSF is similar to U.S. endemic diseases and foreign animal diseases. Diagnostic testing is required to confirm FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 7

18 CSF. Several OIE-approved tests are available; preferred diagnostic samples include serum (for live animals) or tissues such as tonsil and ileum (for dead animals). 45 An overview of CSF diagnostic methods, as described in the OIE Manual of Diagnostic Tests and Vaccines for Terrestrial Animals is given below. 4.1 Detecting Infected Animals by Identifying Virus, Nucleic Acids, or Antigen Identification of the Virus Virus Isolation Virus isolation is a highly sensitive method of CSF diagnosis. It is the test of choice to confirm CSFV infection; however, in an outbreak situation, virus isolation may be too slow and labor intensive to test large numbers of samples. 28 The tonsil is preferred for testing, although spleen, kidney, ileum, and lymph nodes can also be used. 45 In live animals, blood, plasma, and tonsil scrapings can be tested. 68 PK-15 cells or other porcine cells lines can be used, though they must be free of other pestiviruses and pestivirus antibodies. Viral growth does not cause a cytopathic effect. Viral antigen must be detected via the fluorescent antibody test (FAT) after 24 to 72 hours, or in 4- to 5-day-old cultures by immunoperoxidase staining (described in section ). 45 Samples for virus isolation should be refrigerated but not frozen; they should be kept cold during shipment to the laboratory Reverse Transcription Polymerase Chain Reaction (RT-PCR) RT-PCR is a fast and sensitive diagnostic method that can detect CSF nucleic acids in pigs throughout the course of disease. RT-PCR can also be used to detect CSFV in tissues that are autolyzed. 70 It is accepted for CSF testing by the EU and other nations for screening and confirmation of suspected cases of disease. 45,70 Because false positive may occur, primary outbreaks must be confirmed by other tests. RT- PCR protocols, both standard and real-time, have been widely published and are also available from the OIE Reference Laboratories for CSF. Blood samples from live pigs or tissues samples, including tonsil, spleen, ileum and lymph node, collected during necropsy can be tested 28 with results in 48 hours or less. 70 RT-PCR assays can differentiate CSFV from ruminant pestiviruses. 45 Novel pestiviruses that infect pigs, known as atypical porcine pestiviruses, have recently been characterized in Australia, 71 the U.S., 17,18 Germany, 19,20 the Netherlands, 21 Austria, 22,24 and China. 23,25 Concerns about cross-reactivity in genome detection between CSF and atypical porcine pestiviruses have been raised. However, limited data indicate that RT-PCR assays used for CSF diagnosis cannot detect the genomes of atypical porcine pestiviruses Real Time RT-PCR (rrt-pcr) rrt-pcr, also known as quantitative RT-PCR (qrt-pcr), is a variation where nucleic acids are detected as they are being amplified (i.e., in real time ) by an automated process. This makes rrt-pcr a more rapid test compared to standard RT-PCR, and results can be available within 2 hours after the samples are prepared. rrt-pcr can be used when confirming a test result or for surveillance purposes. 68 Tonsil scrapings, tonsil, spleen, lymph node, blood, and nasal swabs can be tested by qrt-pcr. 73 A protocol published by Hoffmann et al. 74 is widely in use according to the OIE. 45 There are a number of other rrt-pcr systems as reported by Blome et al., 26 including those that allow differentiation between vaccine and field virus strains. Methods that involve sample pooling must be validated in individual laboratories, and quality control is essential to prevent contamination that may cause false positives. qrt- PCR has gradually been replacing other antigen detection methods, 26 although virus isolation will remain necessary for further testing. 68 FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 8

19 Genetic Sequencing Following amplification by RT-PCR, sequence data can be obtained in order to compare the genomes of different CSFV isolates. This is particularly important for primary outbreaks, where CSFV isolates should be sent to an OIE Reference Laboratory for analysis. According to the OIE, 45 the regions that are sequenced most frequently are the 5 -nontranslated region (5 NTR) and the E2 major glycoprotein gene Identification of Viral Antigen Fluorescent Antibody Test (FAT) CSFV antigen can be rapidly detected using FAT. This test uses frozen sections of tonsils, spleen, kidney, lymph nodes or distal portions of the ileum which are stained directly with an anti-csf immunoglobulin conjugated to a fluorescence marker, or indirectly with a fluorescent conjugate and examined by fluorescence microscopy. 45 Detecting CSF virus in the tonsil is more likely 2 to 15 days post infection when animals are showing clinical signs of CSF compared to animals that have been infected for a longer period of time. 49,75 Testing the ileum will provide more accurate test results for subacute and chronic cases. 28 If a FAT result is negative and CSF is still suspected, RT-PCR or virus isolation in cell culture should be attempted. 45 Only laboratories that perform the FAT on a regular basis should be used to minimize the risk of false positives. 45 Vaccine administration and infection with ruminant pestiviruses may affect FAT results in some cases. Administration of the live attenuated virus vaccine may cause pigs to test positive on the FAT for 2 weeks following vaccination. Ruminant pestiviruses can also interfere with CSFV testing causing false-positive FAT reactions. Pigs infected with ruminant pestiviruses from congenital infections can have the same clinical signs and lesions as pigs with chronic CSF. 76,77 To differentiate CSFV infection from infection with ruminant pestiviruses, animals can be tested for neutralizing antibodies to the virus. 45 It may also be possible for atypical porcine pestivirus infection to confound FAT results Immunoperoxidase Staining Immunoperoxidase staining is a test that can be used to differentiate field strains of CSFV, vaccine strains of CSFV, and other pestiviruses including those from ruminants. Immunoperoxidase staining has been used in the investigation of an atypical porcine pestivirus (Bungowannah virus) in Australia. 71 The test uses monoclonal antibodies (MAbs) that are tagged with an enzyme; a chemical reaction occurs following CSFV antigen and antibody binding and produces a colored product. 68 In a given geographic region, MAbs must be specific to the CSF strains in circulation and vaccine strains being used, if any Antigen-Capture Enzyme Linked Immunosorbent Assay (ELISA) Antigen-capture ELISA is a good tool for early diagnosis of CSFV at the herd level; however, sensitivity is low and the assay is not appropriate for CSFV surveillance purposes or to detect CSFV in individual animals. 28 Blood, tissues, plasma, or serum specimens can be tested. 68 Most commercially available tests detect the CSFV glycoprotein Erns Detecting Infected Animals by Serological Assays Serology is used for diagnosis and surveillance, especially when infection with a CSFV strain of low virulence is suspected. 45 It is also useful in the final phase of CSF eradication when trying to detect positive animals that might remain in a breeding herd. 45 Antibodies cannot be detected until at least 21 days post-infection and can be present for the life of the animal. 45,78 Congenitally infected pigs are immunotolerant and do not produce antibodies that are detectable via serology. 36 FAD PReP/NAHEMS Guidelines: Appendix B: Vaccination for Classical Swine Fever (2017) 9